A substantial body of evidence suggests that nicotine adversely affects cerebral blood flow and peripheral thrombus formation by inducing breakdown of the blood-brain barrier and alterations in the cerebrovascular endothelium. Some of the alterations in brain endothelial phenotype and function are denoted as proinflammatory, indicating that nicotine may regulate expression of some proinflammatory mediators such as cytokines and adhesion molecules. Our preliminary data support this observation and indicate that chronic exposure to nicotine (14 days) has a profound effect on postischemic injury (2-fold increase in infarct size) and on the progression of the inflammatory response. Particularly, administration of nicotine markedly increased leukocyte migration into postischemic mouse brain and also induced high expression of chemokine CCL2. That chemokine is a major factor orchestrating the immune response and it plays a pivotal role in leukocyte recruitment during postichemic inflammation. To further explore this issue, we propose a research plan directed to test the hypothesis: "Nicotine augments postischemic inflammatory response in the brain by inducing a prolonged increase in expression of chemokines." Specifically, we will determine whether nicotine and/or cotonine exposure can induce or maintain the inflammatory response of brain endothelial cells through the stimulation of CCL2 expression in these cells under in vitro and in vivo basal (i.e., normal oxygen and glucose supply) and ischemia/reperfusion (I/R) conditions (specific aim 1) and to establish the components of the signaling pathway (transcription factor NF:B and Rho/Rho kinase pathway) that underlie the upregulation of CCL2 expression in mouse brain endothelial cells by nicotine during postischemic injury (specific aim 2). To accomplish our goal, we will conduct both in vitro and in vivo experiments. For in vitro experiments, brain microvessel endothelial cell cultures will be subjected to combined oxygen glucose deprivation followed by recovery (in vitro model of I/R). In vivo experiments will be performed on a mouse model of brain I/R injury (transient middle cerebral artery occlusion). Collectively, these experiments will serve to illuminate, for the first time, how nicotine can regulate postischemic inflammatory response. PUBLIC HEALTH RELEVANCE: Cigarette smoking is widely recognized as a major modifiable risk factor for stroke and it is causally associated with a worse stroke outcome. The purpose of this study is to provide a better understanding of the relationship between cigarette smoking and stroke and this may lead to the design of a rational therapeutic approach to stroke in smokers.